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Available from Riken BioResource Center.

Plasmid & Protein Purification Database -Purification-

Species  
Locus tag ST2055
Products  
Plasmid ID R254
Vector  
Purification Lot ST2055_5
Purified Date 2005.6.20-6.30
Protein conc. (mg/ml) 11.0
Total volume (ml) 0.89
Total protein (mg) 9.79

Culture Lot:

date host cell scale label weight of bacteira (g)
        60.0

Cell Lysis and Heat Treatment:

60.0 g of E. coli.
130 ml of 20mM Tris-HCl (pH8.0), 5mM B-ME, 50mM NaCl.
Sonication, output 8, duty 50/2 min, 5 times.
Heat 10 min, 75 deg C.
Centrifuge at 100,000 rcf, 60 min.
120 ml of supernatant.
 
 
 

Purification Procedure:

Step Column Buffer Gradient 1 Gradient 2 FlowRate (ml/min) Eluted
1 Resource Phe 6ml 20mM Tris-HCl(pH8.0),5mM B-ME,1.2M AS 1.2M AS 0M AS 6 Flow through
2 Resource Phe 6ml 20mM Tris-HCl(pH8.0),5mM B-ME,1.8M AS 1.8M AS 0M AS 6 1.7M AS
3 Resource Q 6 ml 20mM Tris-HCl(pH8.0),5mM B-ME 0M NaCl 2M NaCl 4 Flow through
4 HiPrep16/10 Heparin FF 20mM Tris-HCl(pH8.0),5mM B-ME 0M NaCl 2M NaCl 2 Flow through
5 HiLoad 16/60 Superdex 75 20mM Tris-HCl(pH8.0),5mM B-ME,150mM NaCl     1  
6            
7            
8            
9            

Comment:

 

2011.10.20

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